Dermatomycosis vaccine

ABSTRACT

The present invention relates to the preparation of universal inactivated vaccines and their use in preparing compositions for the prophylaxis and therapy of dermatomycosis. Vaccines according to the present invention have the advantage of conferring immunity against all important causes of dermatomycosis in animals and are characterized by stable immunogenic properties, easy preparation, high content of microconidia and lack of side reactions in animals.

RELATED APPLICATIONS

[0001] The present application is a continuation of U.S. patentapplication Ser. No. 10/085,703, filed Feb. 8, 2002, which is acontinuation of U.S. patent application Ser. No. 09/256,915, filed Feb.24, 1999, which is a continuation of U.S. patent application Ser. No.08/568,063, filed Dec. 6, 1995, now abandoned, which is a continuationof U.S. patent application Ser. No. 08/281,380, filed Jul. 26, 1994, nowabandoned, which is a continuation of U.S. patent application Ser. No.08/081,299, filed Aug. 11, 1993, now abandoned, which claims priorityunder 35 USC § 119 to Russian Federation application Serial No.50068611307308, filed Oct. 21, 1991; and claims priority benefit of allthe above-listed applications.

BACKGROUND

[0002] This invention relates to the preparation of vaccines and theiruse in preparing compositions for specifically preventing and treatingdermatomycosis.

[0003] Dermatomycoses in animals are anthropozoonotic diseases of theskin and related tissue. Clinical symptoms are characterized by loss ofhair in the affected area, hyperemia, scaling and asbestos-like scabs.Inflammation is often accompanied by suppuration. Dermatomycoses areoften also characterized by localized infection of the skin.

[0004] Dermatomycoses in animals carry a substantial socioeconomicimpact. Diseased animals required prolonged treatment and can spreadinfection to both animals and humans.

[0005] Up till now, dermatomycoses have been treated using various typesof medication applied locally to affected areas of the skin. Theseincluded the ointments Yam, Yuglon (I) and a number of other ointments,liniments, solutions and other substances containing fungicides andfungistatic agents.

[0006] The disadvantages of such treatments were:

[0007] they were not very effective;

[0008] they required the adoption of quarantine measures anddisinfection of areas where animals were kept (rearing pens, vivaria,farms, zoos, circuses, etc.);

[0009] they required substantial funds to be spent on drug preparationsand veterinary treatment;

[0010] they posed difficulties in immobilizing the animals (for wildanimals held in captivity).

[0011] Later vaccines were developed to treat trichophytosis in cattle(see USSR Patent No. 268593, 1970), fur-bearing animals and rabbits (seeUSSR Patent No. 835446, 1980), camels (see USSR Patent No. 1190574,1985) and others.

[0012] A vaccine had also been developed earlier for the prevention andtreatment of trichophytosis in horses: S-P-I (see USSR Patent No.548947, 1976).

[0013] The S-P-I vaccine contains the vaccinal strain Trichophytonequinum No. 2251/71, deposited with the USSR All-Union State ScientificControl Institute of Veterinary Preparations, which is cultivated inagar/wort for 20-25 days at a temperature of 26-28° C. The fungal massis then lifted from the surface of the nutrient medium, mixed withsterile distilled water and homogenized, and the concentration of cellsis brought to 600-900 million per ml. The homogenate is transferred to aseparate flask and stabilized with a mixture containing 2-8% gelatine(gelatose) and 10-40% sucrose in the ratio 1:1 (+25%), then lyophilized.

[0014] For prophylactic and treatment purposes the vaccine is injectedinto the muscle tissue of the neck area of juvenile and mature horses intwo doses of 1-2 cc, depending on the age of the horse, with an intervalof 10-14 days. For therapeutic use the dosages were doubled.

[0015] Vaccines obtained using this method have the disadvantage thatthey do not provide immunity against microsporiae and trichophytiaecaused by other agents. It has also been noted that the areas where alive vaccine is injected may become a specific focus in which culturesof vaccinal strains may at certain times be produced. Given that somespecies of domestic animals come into frequent contact with humans, theoccurrence of such specific foci in these animals is unacceptable.

DESCRIPTION

[0016] This invention now provides universal inactivated vaccines forthe specific treatment and prevention of dermatomycosis in animals andcorresponding immunogenic fungal strains.

[0017] This aim has been achieved by using the following fungal strainsas vaccinal strains: Trichophyton verrucosum (especially No.VKPGF-931/410), Trichophyton mentagrophytes (especially No.VKPGF-930/1032), Trichophyton equinum (especially No. VKPGF-929/381),Trichophyton sarkisovii (especially No. VKPGF-551/68), Microsporum canis(especially No. VKPGF-928/1393), Microsporum canis var. obesum(especially No. VKPGF-727/1311), Microsporum canis var. distortum(especially No. VKPGF-728/120), Microsporum gypseum (especially No.VKPGF-729/59). Vaccines can be produced by using various combinations ofantigenic material from the above strains together with a suitablecarrier.

[0018] A preferred combination consists of Trichophyton verrucosum No.VKPGF-931/410, Trichophyton mentagrophytes No. VKPGF-930/1032,Trichophyton equinum No. VKPGF-929/381, Microsporum canis No.VKPGF-928/1393, Microsporum canis var. obesum No. VKPGF-727/13 11,Microsporum canis var. distortum No. VKPGF-728/120, Microsporum gypseumNo. VKPGF-729/59, particularly for use in dogs, cats and horses.

[0019] Another preferred combination of vaccine strains consists ofTrichophyton verrucosum No. VKPGF-931/410, Trichophyton mentagrophytesNo. VKPGF-930/1032, Trichophyton sarkisovii No. VKPGF-551/68,particularly for use in cattle.

[0020] The antigenic material may comprise a single antigen of at leastone, and more particularly of all of the above-mentioned dermatophytesor from a plurality of antigens, provided that a sufficient immuneresponse is stimulated to give resistance to a dermatophyte infection.Antigenic material for such a purpose can be prepared using methodsknown from the prior art, e.g., homogenizing the above-mentioneddermatophytes or parts thereof, fractionation of dermatophytepreparations, production of antigenic dermatophyte material byrecombinant DNA technology, etc. It is preferable to use homogenizedculture material having 4 to 120 million, preferably 90 millionmicroconidia.

[0021] Suitable physiologically acceptable carriers for administeringthe vaccines are known from the prior art and may include buffers, gels,microparticles, implantable solids, solutions and other adjuvants.

[0022] To kill off the dermatophytes it is possible to use thiomersal(C₉H₉O₂SNaHg), formaldehyde or 2-propiolactone.

[0023] In order to prepare a vaccine the following procedure may beused, for example.

[0024] Cultures of the strains are homogenized in an aqueous solutioncontaining 0.2 ti 2.0% fermented, hydrolyzed muscle protein (FGM-s), 5to 12% glucose and 0.1 to 1.2% yeast extract. The concentration of themicroconidia is adjusted to 4 to 120 million per milliliter and after 1to 2 days the mixture is inactivated, e.g., with thiomersal(C₉H₉O₂SNaHg) in the ration 1:10,000 to 1:25,000, or with anothersubstance known from the prior art. The resulting suspension is packagedand is ready for use in animals.

[0025] The preparation of the vaccines, the dosage to be given and themethod of administration for prevention and therapeutic treatment areexplained in Examples 1 to 3.

[0026] The invention now makes it possible to prepare an inactivatedvaccine that reduces the probability of reinfection and also implants ahigh degree of immunity. Unlike the known vaccines, the vaccineaccording to the invention in practice gives immunity to all importantcauses of dermatomycosis in animals.

[0027] Briefly, the vaccine according to the invention offers thefollowing advantages:

[0028] in many species of disease-prone animals it establishes immunityafter intramuscular injection,

[0029] it grants immunity against almost all causes of dermatomycosis inanimals,

[0030] it has stable immunogenic properties,

[0031] it is easy to prepare,

[0032] it has a complete set of exo- and endo-antigens of dermatophytecultures and shows no side reactions in animals.

[0033] The vaccine has been successfully tested on over 500 animals ofdifferent species, predominantly in affected regions.

[0034] The strains used to produce the vaccine have been deposited atthe “All-Union Collection of Pathogenic Fungi within the USSR, Ministryof Health Centre for Deep Mycoses” in Leningrad and at the “DSM—DeutscheSammlung von Mikroorganismen und Zellkulturen”, Mascheroder Weg 1B,W-3300 Braunschweig, Germany.

[0035] The following microorganisms were deposited with the DeutscheSammlung von Mikroorganismen und Zellkulturen (DSM) on Oct. 1, 1992 andreceived the following accession numbers:

[0036]T. verrucosum VKPGF-931/410 received accession No. DSM 7277

[0037]T. mentagrophytes VKPGF-930/1032 received accession No. DSM 7279

[0038]T. equinum VKPGF-929/381 received accession No. DSM 7276

[0039]T. sarkisovii VKPGF-551/68 received accession No. DSM 7278

[0040]M. canis VKPGF-928/1393 received accession No. DSM 7281

[0041]M. canis var. obesurn VKPGF-727/1311 received accession No. DSM7280

[0042]M. canis var. distorluin VKPGF-728/120 received accession No. DSM7275

[0043]M. gypseun VKPGF-729/59 received accession No. DSM 7274.

[0044] DSM is located at Macheroder Weg 1B, W-3300 Braunschweig,Germany. “T” is an abbreviation for Trichophyton and “M” is anabbreviation for Microsporum.

[0045] Their characteristics are set out below:

[0046]Trichophyton Verrucosum, No. VKPGF-931/410

[0047] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismeni und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0048] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 410, which wasidentified on a deer in 1978. The strain was identified using theRebell-Taplin key (Rebell, G., Taplin, D.: Dermatophytes, theirrecognition and identification, 1978) and according to Kashkin, P. N.et. al. (Opredelitel patogennykh, toksigenykh vrednykh dlya chelovekagribov, 1979).

[0049] The biological properties of the strain are described in Table 1.

[0050] Strain No. VKPGF-931/410 differs from the epizootic strain in itsfaster growth in nutrient medium, the enormous production ofmicroconidia, lower virulence and the absence of any reaction with itsantigens.

[0051]Trichophyton Mentagrophytes No. VKPGF-930/1032

[0052] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0053] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 1032, which wasfound on a horse in 1985. The strain was identified as described aboveRebel, Taplin, loc. cit. and Kashkin, loc. cit.). The biologicalproperties are described in Table 2.

[0054] Strain No. VKPGF-930/1032 differs from the epizootic strain byits faster growth in nutrient medium, the enormous production ofmicroconidia, its lower virulence and the absence of any reaction withits antigens.

[0055]Trichophyton Equinum No. VKPGF-929/381

[0056] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0057] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 381 which wasfound on a horse in 1986. The strain was identified as describe aboveRebel, Taplin, loc. cit. and Kaslikin, loc. cit.). The biologicalproperties are described in Table 3.

[0058] Strain No. VKPGF-929/381 differs from the epizootic strain by itsfaster growth in nutrient medium, lower virulence and the absence of anyreaction with its antigens.

[0059]Microsporum Canis No. VKPGF-928/1393

[0060] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0061] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 1393 which wasfound on a cat in 1988. The strain was identified as describe aboveRebel, Taplin, loc. cit. and Kashkin, loc. cit.). The biologicalproperties are described in Table 4.

[0062] Strain No. VKPGF-928/1393 differs from the epizootic strain byits faster growth in nutrient medium, its enormous capacity to carryspores, lower virulence and the absence of any reaction with itsantigens.

[0063]Microsporum Canis Var. Obesum No. VKPGF-727/1311

[0064] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0065] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 1311 which wasfound on a tiger in 1986. The strain was identified as describe aboveRebel, Taplin, loc. cit. and Kashkin, loc. cit.). The biologicalproperties are described in Table 5.

[0066] Strain No. VKPGF-727/1311 differs from the epizootic strain byits faster growth in nutrient medium, its enormous capacity to carryspores, lower virulence and the absence of any reaction with itsantigens.

[0067]Microsporum Canis Var. Distortum No. VKPGF-728/120

[0068] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganisrnen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0069] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 120 which wasfound on a black panther in 1987. The strain was identified as describeabove Rebel, Taplin, loc. cit. and Kashkin, loc. cit.). The biologicalproperties are described in Table 6.

[0070] Strain No. VKPGF-728/120 differs from the epizootic strain by itsfaster growth in nutrient medium, its enormnous production ofmicroconidia, its lower virulence and the absence of any reaction withits antigens.

[0071]Microsporum Gypseum No. VKPGF-729/59

[0072] The strain was deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen”, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0073] The strain was obtained by directed selection based on sporeproduction and attenuation of the epizootic Strain No. 59 which wasfound on a horse in 1985. The strain was identified as describe aboveRebel, Taplin, loc. cit. and Kashkin, loc. cit.). The biologicalproperties are described in Table 7.

[0074] Strain No. VKPGF-729/59 differs from the epizootic strain by itsfaster growth in nutrient medium, the enormous production ofmicroconidia, the lower virulence and the absence of any reaction withits antigens. TABLE 1 Properties and characteristics of strain StrainNo. VKPGF-931/410 Epizootic Strain No. 410 Description of culture Mature10-15 day single-spore colony in Mature 25-30 day colony in agar/wort;cream, agar/wort; white, velvety, convex, narrow leathery/velvety,folded, undersurface growing margin undersurface colorless, colorless,colony diameter 9-13 mm colony diameter 10-15 mm Morphologicalcharacteristics Mature 10-15 day culture with septate Mature 25-30 dayculture with septate branching branching hyphae 1-3 μm wide, numerousmycelium 1-3 μm wide, few oval, pyriform, oval, pyriform microconidiameasuring 1.5 cylindrical microconidia measuring 1 to 3 × 3 to 7 μm, to3 × 3 to 5 μm, no macroconidia single elongate irregular shapemacroconidia with 2-5 septates measuring 3 to 5 × 25 to 30 μm, numerousarthrospores in chains 6-8 μm diameter, chlamydospores 10-12 μm diameterPathogenic characteristics Thin necrotic scabs Dense asbestos-likescabs, possible suppuration 12 to 15 days after application of a dose of500-600 19-20 days 25-30 days thousand cells of fungal matter per cm² tothe scarified skin of a rabbit Spontaneous recovery after Reactionresponse No observed changes in clinical state Inflammation at point ofinjection, edema Results of subcutaneous and intramuscular injection ofinactivated corpuscular antigens from cultures Antigen response 20 to 25days after injecting rabbits with corpuscular antigens, antibody titersobserved in blood serum By Passive Hemagglutination Reaction (PHR) 1:320to 1:640 1:320 to 1:640 By Enzyme-linked Immunosorbent Assay (ELISA)1:400 to 1:1600 1:400 to 1:1600 Immunogenic response Establishesimmunity Establishes immunity Immunization of a group of rabbits withinactivated antigens from cultures (repeated at least 5 times)

[0075] TABLE 2 Properties and characteristics of strain Strain No.VKPGF-930/1032 Epizootic Strain No. 1032 Description of culture Mature10-15 day colony in agar/wort; Mature 25-30 day colony in agar/wort;cream, velvety/powdered, flat with white, flat, narrow growing margin,slight flat elevation in center, narrow undersurface reddish-brown,colony growing margin, fringed, undersurface diameter 15-20 mm lightbrown, colony diameter 25-30 mm Morphological characteristics Septate,branching hyphae 1-3 μm Septate, branching straight and spiral wide,numerous pyriform, oval hyphae 1-3 μm wide, round, flattenedmicroconidia measuring 1 to 3 × 2 to 6 μm, pyriform microconidiameasuring 1 to no macroconidia 3 × 2 to 6 μm, few elongate-ovalmacroconidia with 2-5 septates, measuring 2 to 6 × 15 to 25 μmPathogenic characteristics Necrotic scabs Dense, asbestos-like scabs 9to 10 days after application of a dose of 500-600 22-25 days 30-35 daysthousand cells of fungal matter per cm² to the scarified skin of arabbit Spontaneous recovery after Reaction response No observed changesin clinical state Inflammation at point of injection, Results ofsubcutaneous and intramuscular injection of edema inactivatedcorpuscular antigens from cultures Antigen response 20 to 25 days afterinjecting rabbits with corpuscular antigens, antibody titers observed inblood serum By PHR 1:320 to 1:640 1:320 to 1:640 By ELISA 1:400 to1:1600 1:400 to 1:1600 Immunogenic response Establishes immunityEstablishes immunity Immunization of a group of rabbits with inactivatedantigens from cultures (repeated at least 5 times)

[0076] TABLE 3 Properties and characteristics of strain Strain No.VKPGF-929/381 Epizootic Strain No. 381 Description of culture Mature10-15 day colony in agar/wort; Mature 15 day colony in agar/wort; white,velvety/powdery, flat with slight white, velvety, slightly creasedelevation in center, narrow growing center, narrow growing margin,margin, fringed, undersurface light undersurface reddish-brown, brown,colony colony diameter diameter 15-20 mm 13-15 mm Morphologicalcharacteristics Septate, branching hyphae 1-3 μm Septate, branchinghyphae with coil wide, numerous oval pyriform end, 1-4 μm wide, fewoval, microconidia measuring 2 to 3 × 3 to 6 μm, pyriform microconidiameasuring no macroconidia 2 to 3 × 3 to 7 μm, lobar macroconidiameasuring 4 to 7 × 15 to 25 μm Pathogenic characteristics Necrotic scabsAsbestos-like scabs 10 to 12 days after application of a dose of 500-600thousand cells 20-22 days 25-30 days of fungal matter per cm² to thescarified skin of a rabbit Spontaneous recovery after Reaction responseNo observed changes in clinical state Inflammation at point ofinjection, Results of subcutaneous and intramuscular injection ofinactivated edema corpuscular antigens from cultures Antigen response 20to 25 days after injecting rabbits with corpuscular antigens, antibodytiters observed in blood serum By PHR 1:320 to 1:640 1:320 to 1:640 ByELISA 1:800 to 1:1600 1:800 to 1:1600 Immunogenic response Establishesimmunity Establishes immunity Immunization of a group of rabbits withinactivated antigens from cultures (repeated at least 5 times)

[0077] TABLE 4 Properties and characteristics of strain Strain No.VKPGF-928/1393 Epizootic Strain No. 1393 Description of culture Mature10-15 day colony in agar/wort; Mature 15 day colony in agar/wort; white,fluffy, convex, narrow growing greyish-beige, arachnoid, powdery inmargin, arachnoid, undersurface brown, center, growing margin fringed,colony diameter 30-35 mm undersurface yellowish, colony diameter 20-25mm Morphological characteristics Septate, branching hyphae 1-4 μm wide,Septate, branching hyphae 2 to 6 μm numerous pyriform, cylindrical wide,few pyriform, cylindrical microconidia, few fusiform microconidiameasuring 1 to 3 × 3 to 7 μm, macroconidia with 3-11 septates, numerousfusiform macroconidia measuring 10 to 20 × 40 to 75 μm with 3-11septates, measuring 10 to 20 × 45 to 85 μm Pathogenic characteristicsNecrotic scabs Dense, asbestos-like scabs 9 to 11 days after applicationof a dose of 500-600 thousand 20-24 days 25-45 days cells of fungalmatter per cm² to the scarified skin of a rabbit Spontaneous recoveryafter Reaction response No observed changes in clinical state Edema andinflammation at point of Results of subcutaneous and intramuscularinjection of injection inactivated corpuscular antigens from culturesAntigen response 20 to 25 days after injecting rabbits with corpuscularantigens, antibody titers observed in blood serum By PHR 1:320 to 1:6401:320 to 1:640 By ELISA 1:400 to 1:1600 1:400 to 1:1600 Immunogenicresponse Establishes immunity Establishes immunity Immunization of agroup of rabbits with inactivated antigens from cultures (repeated atleast 5 times)

[0078] TABLE 5 Properties and characteristics of strain Strain No.VKPGF-727/1311 Epizootic Strain No. 1311 Description of culture Mature10-15 day colony in agar/wort; Mature 15 day colony in agar/wort; white,fluffy, flat with a denser central dome- greyish, fasciculate/arachnoidwith like elevation, narrow growing margin, pieces of cottony whitemycelium, fringed, undersurface colorless with brown growing marginfringed, undersurface center, colony diameter 30-35 mm brownish, colonydiameter 23-28 mm Morphological characteristics Septate, branchinghyphae 1-3 μm Septate, branching hyphae 1-5 μm wide, numerous pyriform,oval and wide, few oval, cylindrical microconidia cylindricalmicroconidia measuring 1 to measuring 1 to 3 × 3 to 8 μm, numerous 3 × 3to 7 μm, few short, elliptical, elliptical, fusiform, elongate-oval orfusiform, elongate-oval macroconidia, irregularly-shaped macroconidiawith some irregular shapes, less frequently 2-5 septates, measuring 11to 20 × 25 “beaked”, with 2-5 septates, measuring to 55 μm 11 to 20 × 25to 50 μm Pathogenic characteristics Thin necrotic scabs Dense,asbestos-like scabs 12 to 15 days after application of a dose of 500-600thousand 10-25 days 25-30 days cells of fungal matter per cm² to thescarified skin of a rabbit Spontaneous recovery after Reaction responseNo observed changes in clinical state Inflammation and edema at point ofResults of subcutaneous and intramuscular injection of injectioninactivated corpuscular antigens from cultures Antigen response 20 to 25days after injecting rabbits with corpuscular antigens, antibody titersobserved in blood serum By PHR 1:320 to 1:640 1:320 to 1:640 By ELISA1:800 to 1:1600 1:800 to 1:1600 Immunogenic response Establishesimmunity Establishes immunity Immunization of a group of rabbits withinactivated antigens from cultures (repeated at least 5 times)

[0079] TABLE 6 Properties and characteristics of strain Strain No.VKPGF-728/120 Epizootic Strain No. 120 Description of culture Mature10-15 day colony in agar/wort; Mature 15 day colony in agar/wort; cream,velvety/powdery, button-like light-beige, powdery, umbonate, elevationin center, narrow growing narrow growing margin, undersurface margin,finely-fringed, undersurface brown, colony diameter 18-20 mm light-brownwith dark-brown center, colony diameter 25-30 mm Morphologicalcharacteristics Septate, branching hyphae 1-3 μm Septate, branchinghyphae 1-3 μm wide, numerous pyriform, oval, wide, few pyriform, oval,cylindrical cylindrical microconidia measuring 1 to microconidiameasuring 1 to 3 × 3 to 3 × 3 to 8 μm, few irregular deformed 8 μm,numerous irregular deformed or macroconidia, distorted or fusiformfusiform macroconidia with 2-9 with 2-9 septates, measuring 8 to 20 × 25septates, measuring 8 to 20 × 25 to to 70 μm 80 μm Pathogeniccharacteristics Thin necrotic scabs Asbestos-like scabs 12 to 15 daysafter application of a dose of 500-600 thousand 20-25 days 27-45 dayscells of fungal matter per cm² to the scarified skin of a rabbitSpontaneous recovery after Reaction response No observed changes inclinical state Inflammation and edema at point of Results ofsubcutaneous and intramuscular injection of injection inactivatedcorpuscular antigens from cultures Antigen response 20 to 25 days afterinjecting rabbits with corpuscular antigens, antibody titers observed inblood serum By PHR 1:320 to 1:640 1:320 to 1:640 By ELISA 1:800 to1:1600 1:800 to 1:1600 Immunogenic response Establishes immunityEstablishes immunity Immunization of a group of rabbits with inactivatedantigens from cultures (repeated at least 5 times)

[0080] TABLE 7 Properties and characteristics of strain Strain No.VKPGF-729/59 Epizootic Strain No. 59 Description of culture Mature 10-15day colony in agar/wort; Mature 15 day colony in agar/wort; white,velvety/fluffy, flat with slight cream, velvety/powdery, flat withfluffy elevation in center of colony, flat white mycelium in center,thin growing growing margin, undersurface brownish, margin, undersurfacebrownish, colony colony diameter 25-30 mm diameter 20-22 mmMorphological characteristics Septate, branching hyphae 2-3 μm wide,Septate, branching hyphae 2-5 μm wide, numerous oval, pyriform,cylindrical few oval, pyriform, cylindrical microconidia measuring 2 to4 × 3 to 6 μm, microconidia measuring 2 to 4 × 3 to no or fewmacroconidia, elliptical, 7 μm, numerous elliptical, stretched-ovalelongate-oval shape with 2-5 septates, macroconidia with 2-5 septates,measuring 7 to 15 × 25 to 40 μm measuring 7 to 15 × 25 to 50 μmPathogenic characteristics Thin necrotic scabs Dense, asbestos-likescabs 12 to 15 days after application of a dose of 500-600 thousand20-22 days 25-28 days cells of fungal matter per cm² to the scarifiedskin of a rabbit Spontaneous recovery after Reaction response Noobserved changes in clinical state Inflammation at point of injectionResults of subcutaneous and intramuscular injection of inactivatedcorpuscular antigens from cultures Antigen response 20 to 25 days afterinjecting rabbits with corpuscular antigens, antibody titers observed inblood serum By PHR 1:320 to 1:640 1:320 to 1:640 By ELISA 1:400 to1:1600 1:400 to 1:1600 Immunogenic response Establishes immunityEstablishes immunity Immunization of a group of rabbits with inactivatedantigens from cultures (repeated at least 5 times)

[0081] The vaccine may be prepared using the strain Trichophytonsarkovii, No. 551/68. It is described for example in USSR Patent No.1177972 dated Aug. 08, 1985, to which reference is made in its entirety.

[0082] This strain was also deposited at the DSM—Deutsche Sammlung vonMikroorganismen und Zellkulturen, Mascheroder Weg 1B, W-3300Braunschweig, Germany.

[0083] In particular, the invention relates to the following:

[0084] a dermatomycosis vaccine, characterized in that it containsantigenic material from at least one of the following dermatophytes:

[0085]Trichophyton verrucosum, particularly Trichophyton verrucosumStrain No. VKPGF-931/410 and/or

[0086]Trichophyton mentagrophytes, particularly Trichophytonmentagroplytes Strain No. VKPGF-930/1032 and/or

[0087]Trichophyton sarkisovii, particularly Trichophyton sarkisoviiStrain No. VKPGF-551/68 and/or

[0088]Microsporum canis, particularly Microsporum canis Strain No.VKPGF-928/1393 and/or

[0089]Microsporum canis var. obesum, particularly Microsporum canis var.obesum Strain No. VKPGF-727/1311 and/or

[0090]Microsporum canis var. distortum, particularly Microsporum canisvar. distortui Strain No. VKPGF-728/120 and/or

[0091]Microsporum gypseum, particularly Microsporuin gypseum Strain No.VKPGF-729/59, and a physiologically acceptable carrier.

[0092] a dermatomycosis vaccine, particularly as an agent for treatingdogs, cats and horses, characterized in that it contains antigenicmaterial from the dermatophyte strains Trichophyton verrucosum No.VKPGF-931/410, Trichophyion mentagrophytes No. VKPGF-930/1032,Trichophyton equinum No. VKPGF-929/381, Trichophyton sarkisovii StrainNo. VKPGF-551/68, Microsporum canis No. VKPGF-928/1393, Microsporumcanis var. obesum No. VKPGF-727/1311, Microsporum canis var. distortumNo. VKPGF-728/120, Microsporuin gypseum No. VKPGF-729/59, together witha physiologically acceptable carrier.

[0093] a dermatomycosis vaccine, more particularly as an agent fortreating cattle, characterized in that it contains antigenic materialfrom the dermatophyte strains Trichophyton verrucosum No. VKPGF-931/410,Trichophyton mentagrophytes No. VKPGF-930/1032, Trichophyton equinum No.VKPGF-929/381, Trichophyton sarkisovii Strain No. VKPGF-551/68, togetherwith a physiologically acceptable carrier.

[0094] a dermatomycosis vaccine as described above, characterized inthat it contains 4 to 120 million, preferably 90 million microconidia,

[0095] a dermatomycosis vaccine as described above, characterized inthat it contains thiomersal or formaldehyde or 2-propiolactone asinactivator,

[0096] a dermatomycosis vaccine as described above, characterized inthat the physiologically acceptable carrier used is an aqueous solutioncontaining 0.2 to 2.0 percent weight of fermented, hydrolyzed muscleprotein, 5 to 12 percent weight glucose and 0.1 to 1.2 percent weightyeast extract,

[0097] the dermatophyte strains:

[0098]Trichophyton verrucosum Strain No. VKPGF-931/410,

[0099]Trichophyton mentagrophytes Strain No. VKPGF-930/1032,

[0100]Trichophyton equinum Strain No. VKPGF-929/381,

[0101]Microsporum canis Strain No. VKPGF-928/1393,

[0102]Microsporum canis var. obesum Strain No. VKPGF-727/1311,

[0103]Microsporum canis var. distortum Strain No. VKPGF-728/120, and

[0104]Microsporum gypseum Strain No. VKPGF-729/59.

[0105] a process for preparing a vaccine, characterized in that:

[0106] a. antigenic material is prepared from at least one of thefollowing strains:

[0107]Trichophyton verrucosum Strain No. VKPGF-931/410,

[0108]Trichophyton mentagrophytes Strain No. VKPGF-930/1032,

[0109]Trichophyton sarkovii Strain No. VKPGF-551/68,

[0110]Microsporum canis Strain No. VKPGF-928/1393,

[0111]Microsporum canis var. obesum Strain No. VKPGF-727/1311,

[0112]Microsporum canis van. distorium Strain No. VKPGF-728/120,

[0113]Microsporum gypseum Strain No. VKPGF-729/59, and

[0114] b. the antigenic material is mixed with a physiologicallyacceptable carrier.

[0115] a process as described above, characterized in that an agent,particularly thiomersal, formaldehyde or 2-propiolactone is added toinactivate the dermatophytes.

[0116] The invention is illustrated by means of the Examples thatfollow.

EXAMPLES Example 1

[0117] To produce 1 liter of vaccine, cultures are taken of the strainsVKPGF-931/410, 930/1032, 929/381, 551/68, 928/1393, 727/1311, 728/120,and 729/59 and grown in agar/wort at 26° C. for 15 days. Each culture isgrown in 8 mattress flasks. The fungal mass is then lifted off,homogenized, placed in 200 ml of solution and added to each mixer. Thesolution used is an aqueous solution containing 1% fermented hydrolyzedmuscle protein, 10% glucose and 1% yeast extract. The concentration ofmicroconidia is brought to 90 million per ml of homogenate. After 2days, 125 ml of each culture in suspension is taken and mixed in asingle container. The vaccine may be prepared by mixing together variouscombinations of the given strains.

[0118] To inactivate the homogenate mixture, thiomersal is addeddirectly to the cell suspension in the ration 1:20,000. 50 mg ofthiomersal is added for every liter of homogenate. The cell mixture isallowed to stand at room temperature for 2 days.

[0119] The resulting vaccine is bottled, checked for sterility, safetyand immunogenic properties in accordance with accepted methods, and keptrefrigerated at 4° C.

[0120] Vaccine produced in this manner was used to immunize animals.

[0121] For prophylactic and treatment purposes the vaccine was used inthe following doses (see Table 8):

Example 2

[0122] The vaccine produced by the method described in Example 1wastested on laboratory animals and various other animals for effectivenessin the prevention and treatment of disease. The results are given inTable 9.

Example 3

[0123] The vaccine produced by the method described in Example 1 wasalso used to treat animals suffering from dermatophytiae. The resultsare given in Table 10. TABLE 8 Dosage (ml) Dosage (ml) Animal family AgeSite of injection Prophylactic Treatment Felidae medium/large cats  1-6months Gluteal muscles 2-5 3-6   6 months+ Gluteal muscles 3-7  4-10Small cats  1-5 months Gluteal muscles   1-1.5   1-1.5   5 months+Gluteal muscles 1-2 1-2 Ursidae 1-12 months Gluteal muscles 1-3 3-5  12months+ Gluteal muscles 3-5 5-6 Procyonidae 1-10 months Gluteal muscles0.3-0.5 0.5  10 months+ Gluteal muscles 0.3-0.5 0.5-1.0 Viverridae 1-12months Gluteal muscles 0.3-0.5 0.5  12 months+ Gluteal muscles 0.5-1.00.5-1.0 Hyaenidae 1-12 months Gluteal muscles 1-3 1-3  12 months Glutealmuscles 3-5 5-6 Canidae 1-10 months Gluteal and 0.3-0.5 0.5-1.0  10months+ shoulder muscles 0.3-1.0 0.5-1.0 Equidae 3-12 months Neck area0.3-0.5 0.5-1.0  12 months+ Neck area 0.5 0.5-1.0 Tyropodae  1-6 monthsShoulder and neck 3-5  5-10   6 months+ area 5-8  7-10 Bovidae 1-12months Neck area 3-5  5-10  12 months Neck area 5-8  7-10

[0124] TABLE 9 Type of animals Number Dosage (cm³) Effectiveness Rabbits10 1.0 No symptoms of disease after infection with virulent Dogs 5 0.3cultures of the fungi T. mentagrophytes, T. verrucosum, T. Domestic cats3 1.0 equinum, M. canis, M. gypseum. Horses 5 0.5 No dermatophytiaelinked to the fungi M. canis and T. Ponies 3 0.3 mentagrophytes afterbeing in direct contact with diseased Camels 2 5.0 animals. Bears 2 3.0Leopards 2 4.0 Hyenas 2 2.0 No dermatophytiae linked to the fungi M.canis and T. Servals 2 3.0 mentagrophytes after being in direct contactwith sources of Ocelots 2 2.0 infection. Lions 2 3.0 Tigers 3 7.0 Nasuas3 0.5 Civets 2 1.0 Rabbits 7 1.5 No symptoms of disease after infectionwith virulent Dogs 3 0.5 cultures of the fungi T. sarkisovii and M.gypseum. Domestic cats 3 1.5 Black panthers 2 5.0 No dermatophytiaelinked to the fungi M. canis, T. Tigers 5 7.0 mentagrophytes and T.verrucosum after being in direct Geese 6 3.0 contact with sources ofinfection. Bears 3 1.0 Dogs 8 0.5 Llamas 2 3.0

[0125] TABLE 10 Type of animals Number Dosage (cm³) Effectiveness Blackpanthers 5 7.0 Affected by microsporosis linked to the fungi M. canis.Black panthers 3 4.0 Recovery took place 12-25 days after immunization.Horses 3 1.0 Ponies 2 0.5 Lions 3 10 Tigers 3 10 Dogs 4 0.5 Bear 1 5.0Hyena 1 5.0 Domestic cats 15 1.5 Affected by microsporosis linked to thefungi M. canis. Dogs 5 0.5 Recovery took place 10-20 days afterimmunization. Horses 5 0.7 Black panther 1 6.0 Affected bytrichophytosis linked to the fungi T. Red foxes 4 1.0 mentagrophytes.Recovery took place 12-15 days after Bears 2 5.0 immunization. Mountainsheep 1 7.0 Horses 15 1.0 Affected by microsporosis linked to the fungiM. equinum. Recovery took place 12-20 days after immunization.

[0126] Bibliography

[0127] (1) Aisenberg, A. A., Noskow, A. I., Kolovatsky, P. P.“Primenenie Yuglona v Veterinarii” in Scientific and TechnicalInformation Bulletin of the State and Scientific Control Committee underthe Moldavian Council of Ministers (1958), p. 88.

[0128] (2) USSR Patent No. 548947 (1976).

What is claimed is:
 1. A dermatomycosis vaccine comprising antigenic material from at least one of the following inactivated dermatophyte strains: Trichophyton verrucosum Strain No. VKPGF-931/410 (accession No. DSM 7277), Trichophyton mentagopliytes Strain No. VKPGF-930/1032 (accession No. DSM 7279), Trichophylon equinum Strain No. VKPGF-929/381 (accession No. DSM 7276), Microsporum canis Strain No. VKPGF-928/1393 (accession No. DSM 7281), Microsporum canis var. obesum Strain No. VKPGF-727/1311 (accession No. DSM 7280), Microsporum canis var. distortum Strain No. VKPGF-728/120 (accession No. DSM 7275), and Microsporum gypseum Strain No. VKPGF-729/59 (accession No. DSM 7274).
 2. The vaccine according to claim 1 further comprising antigenic material from Trichophyton sarkisovii Strain No. VKPGF-551/68 (accession No. DSM 7278),
 3. A dermatomycosis vaccine comprising antigenic material from the following inactivated dermatophyte strains: Trichophyton mentagrophytes Strain No. VKPGF-930/1032 (accession No. DSM 7279), Microsporum canis Strain No. VKPGF-928/1393 (accession No. DSM 7281), Microsporum canis var. obesum Strain No. VKPGF-727/1311 (accession No. DSM 7280), Microsporum canis var. distortum Strain No. VKPGF-728/120 (accession No. DSM 7275), and Microsporum gypseum Strain No. VKPGF-729/59 (accession No. DSM 7274).
 4. A dermatomycosis vaccine comprising inactivated dermatophytes, wherein the inactivated dermatophytes consist of: Trichophyton mentagrophytes Strain No. VKPGF-930/1032 (accession No. DSM 7279), Microsporum canis Strain No. VKPGF-928/1393 (accession No. DSM 7281), Microsporum canis var. obesum Strain No. VKPGF-727/1311 (accession No. DSM 7280), Microsporum canis var. distortum Strain No. VKPGF-728/120 (accession No. DSM 7275), and Microsporum gypseum Strain No. VKPGF-729/59 (accession No. DSM 7274).
 5. The vaccine according to claim 1 comprising 4 to 120 million microconidia per ml.
 6. The vaccine according to claim 4 comprising 4 to 120 million microconidia per ml. 